The Establishment Of Key Process And Test Of Antitumor Activity Of RAPC(Recombinant Allophycocyanin) From A Recombinant E.coli

Tang Zhihong (Marine biology)Directed by Prof. Qin SongRecombinant allophycocyanin (rAPC) is a kind of marine active peptide obtained from recombinant technology. The key process of production and antitumor activity of rAPC was studied in the paper.First, the fermentation of recombinant E.coli P1 was studied. The result showed that balanced DO-stat feeding strategy had significant effect on enhancing final cell density at 5L scale. With balanced DO-stat feeding strategy, the expressed rAPC reached to a content of about 3.52g?(L culture broth)-1 that was more 10 folds than it was in general cultivation. At 20L scale, the expression of rAPC in two–steps fermentation could reach 30.24% that was about 50% higher than it was in one-step fermentation. The feasibility of using inexpensive lactose as an inducer to substitute for IPTG in the cultivation of rAPC in E.coli JR1 was also approved in this paper. Secondly, the purification of rAPC was studied. Three steps of purification process of rAPC were established using FPLC. The results showed that the purity of rAPC reached 98.2%.Thirdly, the quality control of rAPC was investigated. The purified rAPC was an acid protein (pI=6.0) and had a Mr value of approximate 60kD. The immunogenicity of rAPC was identical with that of its natural counterpart. The λmax of rAPC was 240nm.Fourthly, antitumor activity of rAPC was studied in this paper. It was showed that rAPC could obviously inhibit S-180 carcinoma and H22 liver cancer. The highest inhibitory rate reached 61.9% and 62.2%. It was also showed that rAPC could enhance immunity function in primary experiment. This paper provides a simple and efficient process for large scale of rAPC preparation and suggests that rAPC can remarkably inhibit the growth of tumor. The work provides the foundation for the drug exploitation of rAPC.